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A genetic linkage map for watermelon derived from a testcross population: ( Citrullus lanatus var. citroides x C. lanatus var. lanatus) x Citrullus colocynthis.

A genetic linkage map was constructed for watermelon using a testcross population [Plant Accession Griffin 14113 ( Citrullus lanatus var. citroides) x New Hampshire Midget (NHM; C. lanatus var. lanatus)] x U.S. Plant Introduction (PI) 386015 ( Citrullus colocynthis). The map contains 141 randomly amplified polymorphic DNA (RAPD) markers produced by 78 primers, 27 inter-simple sequence repeat (ISSR) markers produced by 17 primers, and a sequence-characterized amplified region (SCAR) marker that was previously reported as linked (1.6 cM) to race-1 Fusarium wilt [incited by Fusarium oxysporum Schlechtend.:Fr. f. sp. niveum (E.F.Sm.) W.C. Synder & H.N. Hans] resistance in watermelon. The map consists of 25 linkage groups. Among them are a large linkage group that contains 22 markers covering a mapping distance of 225.6 cM and six large groups each with 10-20 markers covering a mapping distance of 68.8 to 110.8 cM. There are five additional linkage groups consisting of 3-7 markers per group, each covering a mapping distance of 36.5 to 57.2 cM. The 13 remaining linkage groups are small, each consisting of 2-11 markers covering a mapping distance of 3.5-29.9 cM. The entire map covers a total distance of 1,166.2 cM with an average distance of 8.1 cM between two markers. This map is useful for the further development of markers linked to disease resistance and watermelon fruit qualities.

Journal Article↗

In vitro investigation of a standardized dried extract of Citrullus colocynthis on liver toxicity in adult rats.

A standardized extract of Citrullus colocynthis used as an oral natural laxative in folk medicine was tested for its influence on liver function parameters in vitro. Cytochrome P450 (CYP) dependent production of reactive oxygen species (ROS) under the influence of Citrullus colocynthis extract was investigated by means of stimulated lipid peroxidation (LPO), H2O2 formation and amplified chemiluminescence in rat liver microsomes. In rat liver 9000 x g supernatants 4 monooxygenase reactions mediated by different CYP forms were measured. Putative hepatotoxic effects of Citrullus colocynthis extract were measured by means of potassium and GSH concentrations in and LDH leakage from precision-cut rat liver slices. For possible hepatoprotective effects the influence of the extract on carbon tetrachloride-induced changes of these parameters was investigated. Citrullus colocynthis extract in concentrations higher than 10 microg/ml incubation mixture proved to inhibit lipid peroxidation and ROS-production as well as CYP1A-, 2B- and 3A-dependent reactions with typical substrates. In contrast, H2O2 production was not reduced under the influence of the extract, a slight but significant increase was seen. Citrullus colocynthis extract was found to be free of hepatotoxic effects in concentrations up to 100 microg/ml incubation mixture when liver slices were incubated in William's medium E for 22 hours. All viability parameters used were not influenced by the extract of Citrullus colocynthis. Carbon tetrachloride induced hepatotoxicity could not be prevented or alleviated. Moreover, the damage was sometimes enhanced by higher extract concentrations.

Animals↗

Comparative analysis of chloroplast DNA variability in wild and cultivated Citrullus species.

PCR amplification and restriction site analysis of chloroplast (cp) DNA regions was used to detect inter- and intraspecific differences in the genus Citrullus. More than 55 C. lanatus and 15 C. colocynthis accessions collected from diverse geographical areas, C. ecirrhosus and C. rehmii were used. Most of the cpDNA variation within Citrullus was the result of large indels and transitions and transversions. Indels at the ndhA, trnS- trnfM and trnC- trnD regions and several substitutions at restriction enzyme sites can be used to separate C. colocynthis from the other Citrullus species. A nucleotide substitution at a restriction enzyme site at the 3' flanking region of ndhF provided a diagnostic haplotype for C. lanatus var. lanatus, the cultivated watermelon. Similarly, a nucleotide substitution at an intergenic spacer region of the trnC- trnD region resulted in a diagnostic haplotype for citron, C. lanatus var. citroides. Several C. lanatus var. citroides accessions showed the var. lanatus haplotype. C. rehmii showed almost the same haplotype as C. lanatus var. citroides with the exception of a unique insertion at a cpSSR site. Since C. ecirrhosus lacks the derived diagnostic nucleotide substitutions of C. lanatus, it is probably the progenitor of the cultivated watermelon. Intraspecific haplotypes detected within C. colocynthis were associated with geographic origin.

Base Sequence↗

Excess chromium alters uptake and translocation of certain nutrients in citrullus.

Citrullus plants were grown in refined sand with varying levels of chromium to determine their tolerance limit to excess chromium. The plants were maintained in control nutrient solution for 24 days and on the 25th day chromium as dichromate was added at 0.05, 0.1, 0.2, 0.3 and 0.4 mM. A control set of plants was grown in the same nutrient solution without chromium. At chromium levels >0.2 mM plants showed growth depression, with chlorosis and loss of turgor of middle leaves. Affected leaves had narrow lamina; tendrils were thin, short and did not have coiling property. Later chlorosis became severe and changed to necrosis in patches. Petiole along with lamina became wilted, rugged and hung down due to complete loss of water. At lower chromium concentration, (0.05, 0.1 and 0.2 mM) only depression in growth was observed. With increase in chromium concentration of nutrient solution accumulation of chromium in different parts of Citrullus was increased. Increase in concentrations of phosphorus, manganese and decrease in iron, copper, zinc and sulphur were observed in leaves. Toxicity of chromium was greater at 0.2-0.4 mM, compared to lower concentrations. Threshold of toxicity and toxicity of Cr in old leaves were, respectively, 0.9 and 3.9 microg g(-1) dry matter of citrullus.

Chromium↗

Insulinotropic effect of Citrullus colocynthis fruit extracts.

Infusions of Citrullus colocynthis Schrad. (Cucurbitaceae) fruits are traditionally used as antidiabetic medication in Mediterranean countries, but to our knowledge no studies have been undertaken so far to determine the possible mechanisms involved in the antidiabetic properties of the fruit. The present study was designed to investigate whether these fruits possess insulinotropic effects. For this purpose, different extracts of Citrullus colocynthis seed components were obtained: RN II (crude extract), RN VI (hydro-alcoholic extract), RN X (purified extract) and RN XVII (beta-pyrazol-1-ylalanine), the major free amino acid present in the seeds. The insulin secretory effects of these different extracts were evaluated in vitro in the isolated rat pancreas and isolated rat islets in the presence of 8.3 mM glucose. All tested extracts, when perfused for 20 min at 0.1 mg/ml, immediately and significantly stimulated insulin secretion. This effect was transient. In addition, the purified extract (RN X) provoked a clear dose-dependent increase in insulin release from isolated islets. Moreover, a significant and persistant increase in pancreatic flow rate appeared during RN VI, RN X and RN XVII perfusions. In conclusion, our results show that different Citrullus colocynthis seed extracts have an insulinotropic effect which could at least partially account for the antidiabetic activities of these fruits.

Amino Acids↗

An estimation of Citrullus colocynthis toxicity for chicks.

Citrullus colocynthis seed was fed at 2% and 10% of the basal diet to 7-d-old Bovans-type chicks for 6 w. Average body weights and efficiency of feed utilization were markedly depressed in the chicks on 10% Citrullus feed, and the serum activities of LDH, AST and CK and concentrations of total lipid and zinc were significantly increased. The concentration of serum total iron binding capacity was particularly reduced in chicks on 2% Citrullus feed. The concentrations of other serum and blood constituents and of hepatic copper, manganese and zinc were not significantly changes. Lesions seen in the intestines, livers, kidneys and other tissues were fully reversed 4 w after removal from the experimental diet.

Administration, Oral↗

Immunostimulating activity of the hot water-soluble polysaccharide extracts of Anacyclus pyrethrum, Alpinia galanga and Citrullus colocynthis.

Hot water polysaccharide extracts of Anacyclus pyrethrum (L.) Link. (family Compositae) Citrullus colocynthis (L.) Schrad. (family Cucurbitaceae) and Alpinia galanga (L.) Willd. (family Zingiberaceae) were tested for their immunostimulating activity in mice. The fractions from Anacyclus pyrethrum and Alpinia galanga showed a marked stimulating effect on the reticulo-endothelial system (RES) and increased the number of peritoneal exudate cells (PEC), and spleen cells of mice. In this case, the optimum doses were 50 and 25 mg/kg for the two fractions, respectively. On the other hand, the polysaccharide extracts of both Anacyclus pyrethrum and Alpinia galanga markedly enhanced the proliferation of the murine spleen cells in vitro using two tests (in vitro and in vivo effect). The results of the in vivo effect at a doses of 50 and 25 mg/kg, showed a stimulation index better than obtained with the in vitro effect at 50 and 25 microg/ml for Anacyclus pyrethrum and Alpinia galanga, respectively. While the extract of Citrullus colocynthis showed much weaker and variable immunostimulating activity.

Adjuvants, Immunologic↗

Simple sequence repeats in watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai).

Simple sequence repeat length polymorphisms were utilized to examine genetic relatedness among accessions of watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai). A size-fractionated TaqI genomic library was screened for the occurrence of dimer and trimer simple sequence repeats (SSRs). A total of 96 (0.53%) SSR-bearing clones were identified and the inserts from 50 of these were sequenced. The dinucleotide repeats (CT)n and (GA)n accounted for 82% of the SSRs sequenced. PCR primer pairs flanking seven SSR loci were used to amplify SSRs from 32 morphologically variable watermelon genotypes from Africa, Europe, Asia, and Mexico and a single accession of Citrullus colocynthis from Chad. Cluster analysis of SSR length polymorphisms delineated 4 groups at the 25% level of genetic similarity. The largest group contained C. lanatus var. lanatus accessions. The second largest group contained only wild and cultivated "citron"-type or C. lanatus var. citroides accessions. The third group contained an accession tentatively identified as C. lanatus var. lanatus but which perhaps is a hybrid between C. lanatus var. lanatus and C. lanatus var. citroides. The fourth group consisted of a single accession identified as C. colocynthis. "Egusi"-type watermelons from Nigeria grouped with C. lanatus var. lanatus. The use of SSRs for watermelon germplasm characterization and genetic diversity studies is discussed.

DNA, Plant↗

1H and 13C NMR signal assignment of cucurbitacin derivatives from Citrullus colocynthis (L.) Schrader and Ecballium elaterium L. (Cucurbitaceae).

2D NMR-derived 1H and 13C NMR signal assignments of six structurally closely related cucurbitacin derivatives are presented. The investigated 2-O-beta-D-glucopyranosylcucurbitacins I, J, K, and L were obtained from Citrullus colocynthis (L.) Schrader whereas the aglyca cucurbitacin E and I were isolated from Ecballium elaterium L.

Carbon Isotopes↗

Effect of combination of Capsicum frutescens and Citrullus colocynthis on growth, haematological and pathophysiological parameters of rats.

The toxicity of diet containing 10% of Capsicum frutescens or 10% of Citrullus colocynthis fruits or their 1:1 mixture (5% + 5%) to rats treated for 6 weeks was evaluated. Body weight loss, inefficiency of feed utilization, diarrhoea, and enterohepatonephropathy characterized C. colocynthis toxicosis in rats. Despite impairment of rat's growth neither nephropathy nor diarrhoea was detected in rats fed the 10% C. frutescens diet. Feeding the mixture of C. frutescens and C. colocynthis caused more pronounced effects and death of rats. Vital organ lesions accompanied by anaemia and leucopenia were correlated with changes in serum ALP, AST and ALT activities with alterations in concentrations of total protein, albumin, urea and other serum constituents. Serum bilirubin concentration did not change.

Anemia↗

Construction of a linkage map and QTL analysis of horticultural traits for watermelon [Citrullus lanatus (THUNB.) MATSUM & NAKAI] using RAPD, RFLP and ISSR markers.

We have been constructing linkage maps for watermelon ( Citrullus lanatus) on the basis of random amplified polymorphic DNA (RAPD), restriction fragment length polymorphism (RFLP), inter-simple sequence repeats (ISSRs) and isozymes using an F(2) population derived from a crossing between a cultivated inbred line (H-7; C. lanatus) and an African wild form (SA-1; C. lanatus). A total of 120 F(2) plants was used for construction of a linkage map using 477 RAPDs, 53 RFLPs, 23 ISSRs and one isozyme markers. Linkage analysis revealed that 554 loci could be mapped to 11 linkage groups that extended for 2,384 centimorgans (cM). While a BC(1) population [(H-7 x SA-1) x H-7] consisting of 60 individuals was grown and scored for quantitative traits. Another linkage map with a total length of 1,729 cM was constructed in the BC(1) using genetic markers found to segregate in the F(2) population. A QTL analysis was applied by means of interval mapping for locating such agronomic traits as hardness of rind, Brix of flesh juice, flesh color (red and yellow) and rind color. The relative order of markers in the BC(1) map was essentially the same as that on the linkage map in the F(2). A total of five QTLs for four agronomic traits was detected. The QTL for hardness of rind was mapped on group 4. The linkage group 8 contained the QTL for sugar content of the flesh as expressed in Brix of the juice. The QTL for red flesh color was detected on groups 2 and 8. The QTL for rind color mapped on the group 3. The present map and QTL analysis may provide a useful tool for breeders by introducing valuable wild watermelon genes to cultivars.

Chromosome Mapping↗

The expression of the Saccharomyces cerevisiae HAL1 gene increases salt tolerance in transgenic watermelon [Citrullus lanatus (Thunb.) Matsun. & Nakai.].

An optimised Agrobacterium-mediated gene transfer protocol was developed in order to obtain watermelon transgenic plants [Citrullus lanatus (Thunb.) Matsun. & Nakai.]. Transformation efficiencies ranged from 2.8% to 5.3%, depending on the cultivar. The method was applied to obtain genetically engineered watermelon plants expressing the Saccharomyces cerevisiae HAL1 gene related to salt tolerance. In order to enhance its constitutive expression in plants, the HAL1 gene was cloned in a pBiN19 plasmid under control of the 35S promoter with a double enhancer sequence from the cauliflower mosaic virus and the RNA4 leader sequence of the alfalfa mosaic virus. This vector was introduced into Agrobacterium tumefaciens strain LBA4404 for further inoculation of watermelon half-cotyledon explants. The introduction of both the neomycin phosphotransferase II and HAL1 genes was assessed in primary transformants (TG1) by polymerase chain reaction analysis and Southern hybridisation. The expression of the HAL1 gene was determined by Northern analysis, and the diploid level of transgenic plants was confirmed by flow cytometry. The presence of the selectable marker gene in the expected Mendelian ratios was demonstrated in TG2 progenies. The TG2 kanamycin-resistant plantlets elongated better and produced new roots and leaves in culture media supplemented with NaCl compared with the control. Salt tolerance was confirmed in a semi-hydroponic system (EC=6 dS m(-1)) on the basis of the higher growth performance of homozygous TG3 lines with respect to their respective azygous control lines without the transgene. The halotolerance observed confirmed the inheritance of the trait and supports the potential usefulness of the HAL1 gene of S. cerevisiae as a molecular tool for genetic engineering of salt-stress protection in other crop species.

Blotting, Northern↗

Elevated plasma citrulline and arginine due to consumption of Citrullus vulgaris (watermelon).

A 19-month-old girl with developmental delay was found to have moderately elevated plasma citrulline and mildly elevated plasma arginine concentrations. Dietary history revealed that she consumed large quantities of watermelon (Citrullus vulgaris), a fruit containing high free citrulline and arginine concentrations. In order to determine whether the patient's high watermelon intake could account for her elevated plasma citrulline and arginine concentrations, we studied the response of plasma citrulline and arginine to ingestion of watermelon in six healthy adult volunteers. All developed markedly elevated plasma citrulline (mean maximum 593 micromol/L, range 386-1069) and moderately elevated plasma arginine (mean maximum 199 micromol/L, range 128-251). Physicians and laboratory personnel performing metabolic investigations should be aware of watermelon-induced citrullinaemia. Its hallmarks are elevated plasma citrulline, and to a lesser extent arginine, in the absence of orotic or arginosuccinic aciduria or hyperammonaemia. This phenomenon has implications for the management of patients with urea cycle and related disorders.

Adult↗

Growth inhibitory activity of cucurbitacin glucosides isolated from Citrullus colocynthis on human breast cancer cells.

Our aim was to study the effects of cucurbitacin glucosides extracted from Citrullus colocynthis leaves on human breast cancer cell growth. Leaves were extracted, resulting in the identification of cucurbitacin B/E glucosides. The cucurbitacin glucoside combination (1:1) inhibited growth of ER(+) MCF-7 and ER(-) MDA-MB-231 human breast cancer cell lines. Cell-cycle analysis showed that treatment with isolated cucurbitacin glucoside combination resulted in accumulation of cells at the G(2)/M phase of the cell cycle. Treated cells showed rapid reduction in the level of the key protein complex necessary to the regulation of G(2) exit and initiation of mitosis, namely the p34(CDC2)/cyclin B1 complex. cucurbitacin glucoside treatment also caused changes in the overall cell morphology from an elongated form to a round-shaped cell, which indicates that cucurbitacin treatment caused impairment of actin filament organization. This profound morphological change might also influence intracellular signaling by molecules such as PKB, resulting in inhibition in the transmission of survival signals. Reduction in PKB phosphorylation and inhibition of survivin, an anti-apoptosis family member, was observed. The treatment caused elevation in p-STAT3 and in p21(WAF), proven to be a STAT3 positive target in absence of survival signals. Cucurbitacin glucoside treatment also induced apoptosis, as measured by Annexin V/propidium iodide staining and by changes in mitochondrial membrane potential (DeltaPsi) using a fluorescent dye, JC-1. We suggest that cucurbitacin glucosides exhibit pleiotropic effects on cells, causing both cell cycle arrest and apoptosis. These results suggest that cucurbitacin glucosides might have therapeutic value against breast cancer cells.

Blotting, Western↗

Watermelon (Citrullus lanatus) hydroperoxide lyase greatly increases C6 aldehyde formation in transgenic leaves.

Fatty acid hydroperoxide lyase (HL) is the key enzyme for the production of the "green note"compounds, leaf aldehyde [(2E)-hexenal] and leaf alcohol [(3Z)-hexenol], in plant tissues. A cDNA encoding HL was cloned from leaves of watermelon (Citrullus lanatus) and expressed in Nicotiana tabacum. The enzyme is 3 times more active with 13-hydroperoxylinolenic acid than with 13-hydroperoxylinoleic acid. The activity against 9-hydroperoxides of polyunsaturated fatty acids is minimal. Enzyme activity of the watermelon HL in the transgenic leaves was approximately 50 times higher than endogenous HL activity in the wild-type N. tabacum plants. When compared with Arabidopsis HL also expressed in N. tabacum, the highest HL activity is 10 times higher in watermelon HL overexpressing leaves than in Arabidopsis HL overexpressers.

Aldehyde-Lyases↗

Biochemical effects of Citrullus colocynthis in normal and diabetic rats.

Diabetes mellitus is one of the most common endocrine diseases. In UAE many traditional plants such as the Citrullus colocynthis (Handal) are used as antidiabetic remedies. The aim of this study was to examine the effect of the aqueous extract of the seed of C. colocynthis on the biochemical parameters of normal and streptozotocin (STZ)-induced diabetic rats. Diabetes mellitus was induced by a single intraperitoneal (60 mg/kg body wt1) injection of STZ. Normal and diabetic rats were fed with the plant extract daily by oral intubation for 2 weeks. Blood sample were collected at the beginning and end of the experiment for the measurement of biochemical parameters. The plasma level of alanine aminotranferase (ALT), alkaline phosphatase (ALP), aspartate aminotransferase (AST), gamma-glutamyl transferase (GGT), lactic dehydrogenase (LDH) increased significantly after the onset of diabetes. Oral administration of the plant extract reduced the plasma level of AST and LDH significantly. However, the plant extract failed to reduce the increased blood level of GGT and ALP in diabetic rats. Blood urea nitrogen (BUN) increased significantly after the onset of diabetes. No significant difference was observed in the blood creatinine, K+, Na+, Ca2+ and P levels of normal and diabetic rats. The plant extract did not have any effect on BUN level, however, it caused an increase in the level of K+, Na+ in diabetic rats. In conclusion, oral administration of the aqueous extract of the C. colocynthis can ameliorate some of the toxic effects of streptozotocin.

Animals↗

Acetohydroxamate inhibition of the activity of urease from dehusked seeds of water melon (Citrullus vulgaris).

Urease from the seeds of watermelon (Citrullus vulgaris) was purified to apparent homogeneity, using two acetone fractionation steps, heat treatment at 48 degrees C and gel filtration through Sephadex G-200. Effect of acetohydroxamic acid (AHA) on the activity of the homogeneous enzyme preparation (sp. act. 3000 +/- 550U/mg protein) was investigated. AHA exhibited a concentration-dependent inhibition both in the presence and absence of the substrate. The inhibition was uncompetitive and the Ki was 2.5 mM. Binding of AHA with the enzyme was reversible, as 63% activity could be restored by dialysis. Time-dependent inhibition revealed a monophasic inhibition of the activity. Addition of beta-mercaptoethanol (ME) gradually abolished the inhibition. Pre-treatment of native enzyme with 8.0 mM ME for 5 min at 30 degrees C exhibited protection against AHA-induced inhibition. The significance of these observations is discussed.

Citrullus↗